Project Summary Considerable interests in understanding and developing therapeutics for cancer have been to study oncogenic lesion reprogrammed metabolism that is the hallmark of cancer. Gain-of-function hot-spot mutations in the isocitrate dehydrogenase genes (IDH) are among the most common genetic alterations in intrahepatic cholangiocarcinoma (ICC). The IDH mutations lead to production of an oncometabolite 2-hydroxygluatrate that perturbs epigenetics and other cellular processes. However, it was not clear how oncogenic IDH1 mutations alter metabolism that could underlie novel vulnerabilities in ICC. To uncover novel insights in IDH1 mutant ICC, we have established and characterized an IDH1 mutant ICC mouse model (GEMM), as well as patient derived models for in vivo disease biology. Leveraging these models, we demonstrate that mutant IDH1 reprograms metabolism including suppression of mitochondrial function and selective hinderance of de novo pyrimidine synthesis, which underlie novel metabolic vulnerability. Coherently, we identified from large-scale screens selective and potent chemical and genetic vulnerabilities of IDH1 mutant cells that impinge on nucleotide metabolism. As such, an important scientific goal, and that of this NIH Pathway to Independence, are to further understand cellular and physiological basis underpinning the crosstalk between reprogrammed metabolism and vulnerabilities for future therapy development. I propose an innovative research program combining cutting-edge metabolomics, proteomics, as well as classic biochemistry, genetics and chemical biology approaches to obtain mechanistic and translational insights in the novel metabolic vulnerabilities of IDH1 mutant ICC using my human and GEMM models. I hypothesize that oncogenic IDH1 mutations lead to reprogrammed nucleotide synthesis that can be leveraged upon to target IDH1 mutant ICC. I will focus on three specific aims: 1) understanding the cellular mechanisms of mutant IDH1 reprogrammed pyrimidine synthesis and its genetic vulnerability; 2) elucidating how pharmacologic modulation of nucleotide synthesis disrupts DNA replication and accumulates DNA damage underlying the hypersensitivity of IDH1 mutant cells; and 3) identifying in vivo determinants of IDH1 mutant ICC sensitivity to drug treatments. Dr. Nabeel Bardeesy's laboratory and Massachusetts General Hospital Cancer Center provide an ideal training environment for the proposed research. I will avail the outstanding mentorships with a spectrum of expertise in metabolism, DNA damage, chemical biology, proteomic analysis, and clinical oncology. Thus, I will acquire necessary trainings in DNA damage response pathways, quantitative proteomics and pre-clinical compound characterizations for mechanistic and translational research during the mentored K99 phase. The Pathway to Independence Award will enable me to expand my scientific and technical repertoire and develop a hypothesis-driven research program, with which I will build an integrative and translational research platform to perform cancer metabolism research independently in my own laboratory.